Paraoxonase (PON1) is an A-esterase that has been found to display a substrate-dependent polymorphism in humans. Determination of PON1 status (which includes both PON1/192 genotype and level of expression) of an individual may allow the use of this enzyme as a biomarker of susceptibility. Because PON1 detoxifies a number of organophosphates and metabolizes bioactive oxidized lipids, studies of PON1 have a direct impact on the susceptibility on pesticide poisoning to cardiovascular diseases, particularly atherosclerosis. Furthermore, the emerging role of PON1 as an "antioxidant" protein may have importance in certain neuro- degenerative diseases such as Parkinson's disease. The general aim of this proposal is to carry out a series of experiments that would add important knowledge on PON1 function and modulation, thus increasing confidence in its use as a reliable marker of susceptibility by better defining variables that may effect its use in large epidemiological studies. Specific aims are: 1. To assess the effects of environmental chemicals on PON1 activity. We have been accumulating convincing evidence suggesting that the level of expression of PON1 is as important as the genotype. At least 13-fold differences in enzyme activity exist within a single PO11 genotype. Modulation of PON1 activity by environmental chemicals may, therefore, represent an important of an individual's PON1 status. We will focus on a number of metals, present in hazardous waste sites, and will measure their ability to interfere with PON1 activity in vitro and in vivo. By using purified Arg/192 or Gln/192 PON1, we will also determine whether either genotype displays higher sensitivity to inhibition by these environmental compounds. 2. To identify additional substrates of PON1. We have shown that the effect of the polymorphism is "reversed" for some organophosphate substrates. Our first objective is, therefore, to assess that the effect of the polymorphism is "reversed" for some organophosphate substrates. A second objective is to identify additional non-toxic substrates hydrolyzing ability of PON1 on a number of organophosphates. A second objective is to identify additional non- toxic substrates, as those tested so far, such as phenylacetate, do not show any polymorphism. Identification of non-toxic, polymorphic substrates, will be extremely useful for a high throughput assay to be used in large epidemiological studies and in the clinical setting.3. To assess the role of PON1 genotypes and levels of expression influencing the toxicity and toxicokinetics of organophosphates and their oxons. For this purpose we will focus on diazinon and diazinon-oxon, and their toxicokinetics will be studied using a physiologically-based kinetic (PBK) model. The effects of PON1 polymorphism will be evaluated by fitting the overall rates of diazinon and metabolite metabolism in the PBK model to the concentration-time data from each mouse from each mouse genotype (PON1+/+; PON =/-; PON1 -/-; PON -/- Arg/192; PON1 -/- Gln/192; the latter two are transgenic mice only expressing one or the other of the human alleles). The resulting values or peak concentrations, areas under the plasma concentration-time curves, clearances, and half-lives will be compared in order to quantitatively discern effects of genotype on toxicokinetics. In addition, various toxicodynamics models will be used to probe for relationships between blood and tissue concentrations of diazinon and diazinon-oxon, and the degree of acetylcholinesterase inhibition. 4.To investigate whether PON1 status is a risk factor in the development of Parkinson's disease. Several epidemiological studies have suggested that exposure to pesticides may be a risk factor for Parkinson's disease, and a recent study suggests that the PON1/Q192 genotype may be a risk factor for the disease. We propose to confirm and expand this preliminary observation by determining PON1 status in groups of control and Parkinson's disease patients that are part of two other projects in this Superfund program.